Isolation of Auxin Signal Transduction Mutants in Arabidopsis

拟南芥中生长素信号转导突变体的分离

• 批准号: 0090996
• 负责人: Jane Murfett
• 金额: $ 33万
• 依托单位: University of Missouri-Columbia
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-04-15 至 2005-03-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0090996&HistoricalAwards=false
• 关键词: Isolation; Auxin; Signal; Transduction; Mutants

The plant hormone auxin is involved in many aspects of growth and development, yet little is known about exactly how this hormone causes changes within plant cells. It is known that when plant cells are exposed to auxin, certain genes are activated: without auxin, they are not expressed (i.e., their protein products are not made), and with auxin they rapidly become expressed at high levels. It is also known that specific DNA sequences within the promoters of these genes (i.e, the parts of the genes that control when and where the genes are expressed) are needed for this response to auxin. An unknown number of molecules are involved in recognizing auxin after it enters the plant cell, and causing these genes to be activated. The goal of this project is to identify some of the molecules involved in this "auxin signal transduction" process. The genetic approach taken in this project involves making mutants in the model plant Arabidopsis that show aberrant expression of auxin-responsive genes. Towards this end, transgenic plants were generated that carried engineered selectable marker genes controlled by auxin responsive promoters. The transgenic plants were mutagenized, then seeds from these plants were grown under conditions that allowed for the selection of plants that showed aberrant expression of the selectable marker genes. One of the selectable markers used was a gene that confers resistance to the antibiotic hygromycin. Non-mutant, transgenic plants were killed by the antibiotic, because they did not contain enough auxin to switch on the resistance gene. However, a few mutant plants were able to grow, because the resistance gene was expressed more highly in them than in non-mutant plants. The other selectable marker used was a gene encoding the enzyme alcohol dehydrogenase (ADH). ADH converts allyl alcohol to a toxic substance, acrolein. Non-mutant transgenic plants that were treated first with auxin, then with allyl alcohol, were killed because the ADH gene was switched on by the auxin treatment. However a few mutant plants survived the treatments because the ADH gene was expressed at lower levels in them. One hypothesis is that these mutant lines contain alterations in the molecules involved in responding to auxin. For some of the mutants, this hypothesis has been tested by seeing if other auxin responsive genes, in addition to the selectable marker genes, show altered expression in the mutants. One goal of thsi project is to continue identifying new mutants that show altered expression of auxin-responsive genes. A few of these mutants will be analyzed in detail, and the mutant genes will be cloned in order to discover what molecules they encode. This work has great potential to enhance our understanding of both plant hormone biology, and signal transduction processes in plants.

植物激素生长素参与生长和发育的许多方面，但人们对这种激素如何引起植物细胞内的变化知之甚少。 已知当植物细胞暴露于生长素时，某些基因被激活：没有生长素，它们不表达（即，它们的蛋白质产物不会被制造出来），并且在生长素的作用下，它们迅速以高水平表达。 我们还知道，这些基因启动子内的特定DNA序列（即控制基因表达的时间和位置的基因部分）是对生长素的这种反应所必需的。 未知数量的分子参与识别生长素进入植物细胞后，并导致这些基因被激活。 这个项目的目标是确定一些参与“生长素信号转导”过程的分子。 该项目中采用的遗传方法涉及在模式植物拟南芥中制造突变体，这些突变体显示生长素响应基因的异常表达。 为此，产生了携带由生长素应答启动子控制的工程化选择标记基因的转基因植物。 将转基因植物诱变，然后在允许选择显示可选择标记基因的异常表达的植物的条件下生长来自这些植物的种子。 所用的选择标记之一是赋予对抗生素潮霉素抗性的基因。 非突变的转基因植物被抗生素杀死，因为它们没有足够的生长素来启动抗性基因。 然而，少数突变体植物能够生长，因为抗性基因在它们中的表达比在非突变体植物中更高。 使用的另一个选择标记是编码乙醇脱氢酶（ADH）的基因。 ADH将烯丙醇转化为有毒物质丙烯醛。 首先用生长素处理，然后用烯丙醇处理的非突变转基因植物被杀死，因为ADH基因被生长素处理打开。 然而，少数突变体植物在处理中存活，因为ADH基因在它们中的表达水平较低。 一种假设是，这些突变株系包含对生长素反应的分子改变。 对于一些突变体，已经通过观察除了选择标记基因之外的其他生长素响应基因是否在突变体中显示改变的表达来测试该假设。 这个项目的一个目标是继续鉴定新的突变体，这些突变体显示出生长素反应基因表达的改变。 将对其中的一些突变体进行详细分析，并克隆突变基因，以发现它们编码的分子。 这项工作有很大的潜力，以提高我们对植物激素生物学和植物信号转导过程的理解。